ABSTRACT Additive manufacturing is transforming how microfluidic devices are prototyped and fabricated. Among various 3D printing methods, stereolithography (SLA) has become a dominant technique for microfluidics due to its high resolution and design flexibility, with widespread use in lab‐on‐a‐chip applications. However, intrinsic limitations of SLA printing, such as challenges related to multi‐material integration and microstructure fabrication in enclosed channels, continue to hinder the development of more complex microsystems, especially for analytical separation and tissue engineering applications. In this paper, we present a multiphase flow‐assisted in situ 3D printing method to address these challenges, developed based on our previously reported in situ 3D polymerization (IS‐3DP) concept. Our method utilizes an aqueous two‐phase system (ATPS) to generate sequential printing layers through controlled fluidic confinement and integrates an image‐guided alignment system to enable precise projection of printing patterns in microchannels. We demonstrate that viscosity tuning of the ATPS printing and blocking phases enables dynamic control of layer thickness, allowing customized and adaptive design of the 3D structure slicing. The image‐guided alignment system employs a homography transformation mechanism to map the projection and printing planes via image feedback, providing real‐time mask alignment with microchannel geometries. We characterize the mapping accuracy and projection fidelity and demonstrate the capability of this method by direct in‐channel fabrication of complex 3D microstructures such as pyramids, cuboids, bridge‐like void structures, as well as multi‐material patterns. We envision the multiphase flow‐assisted in situ 3D printing to offer a versatile tool for spatially controlled, high‐fidelity, and multi‐material microfabrication within confined microchannels in novel lab‐on‐a‐chip applications.
Ramirez‐Alvarado et al. (2026) studied this question.